Engineered Mutation 'B6.Cg-Ndor1Tg(UBC-cre/ERT2)1Ejb/2J'
Mutation Name B6.Cg-Ndor1Tg(UBC-cre/ERT2)1Ejb/2J
Common Name UBC-Cre
Mutation Type Other Transgenic
Strain of Origin
Gene Symbol
Gene Name
Synonyms
Accession Numbers
Alleles Listed at MGI
Chromosome
Chromosomal Location
Repository

JAX strain name: B6.Cg-Ndor1Tg(UBC-cre/ERT2)1Ejb/2J

Stock No: 008085
Phenotypic Description

Mice hemizygous for this Cre-ERT2 transgene are viable and fertile. Mice from this founder line have strong tamoxifen-inducible cre activity in all reported tissue types. The Cre-ERT2 fusion protein consists of Cre recombinase fused to a triple mutant form of the human estrogen receptor; which does not bind its natural ligand (17β-estradiol) at physiological concentrations but will bind the synthetic estrogen receptor ligands 4-hydroxytamoxifen (OHT) and, with lesser sensitivity, ICI 182780. Restricted to the cytoplasm, Cre-ERT2 can only gain access to the nuclear compartment after exposure to OHT. To counteract the mixed estrogen agonist effects of tamoxifen injections, which can result in late fetal abortions in pregnant mice, progesterone may be coadministered. When these Cre-ERT2 mice are bred with mice containing a loxP-flanked sequence of interest, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the flanked sequences in widespread cells/tissues.
Nature of Mutation

These transgenic mice express a Cre-ERT2 fusion gene under the control of the human ubiquitin C (UBC) promoter.
Expression/Localization

When these Cre-ERT2 mice are bred with mice containing a loxP-flanked sequence of interest, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the flanked sequences in widespread cells/tissues.
Genotyping

(protocol for Dr. Beutler's lab_optimized by Baifang Qin)

NOTES: This assay will NOT distinguish hemizygous from homozygous transgenic animals.

Expected Results:

Transgene =  ~475 bp
Internal positive control = 324 bp

Primers:

PRIMER 5' LABEL SEQUENCE 5' → 3' 3' LABEL PRIMER TYPE REACTION NOTE
25285   GAC GTC ACC CGT TCT GTT G   Transgene Forward A UBC
oIMR7338   CTA GGC CAC AGA ATT GAA AGA TCT   Internal Positive Control Forward A  
oIMR7339   GTA GGT GGA AAT TCT AGC ATC ATC C   Internal Positive Control Reverse A  
oIMR9074   AGG CAA ATT TTG GTG TAC GG   Transgene Reverse A

Cre

Reaction:

 

Components

Component Volume (μL)
JumpStart Mix 15
5 uM F/R 1
Water 11
gDNA 3
TOTAL 30

Cycling:

STEP TEMP °C TIME NOTE
1 94.0 --  
2 94.0 --  
3 65.0 -- -0.5 C per cycle decrease
4 68.0 --  
5   -- repeat steps 2-4 for 10 cycles (Touchdown)
6 94.0 --  
7 60.0 --  
8 72.0 --  
9   -- repeat steps 6-8 for 21 cycles
10 72.0 --  
11 10.0 -- hold

Results can be detected by gel electrophoresis on a 2.0% agarose gel
Notes

MGI: J:123200
References

Ruzankina Y; Pinzon-Guzman C; Asare A; Ong T; Pontano L; Cotsarelis G; Zediak VP; Velez M; Bhandoola A; Brown EJ. 2007. Deletion of the developmentally essential gene ATR in adult mice leads to age-related phenotypes and stem cell loss. Cell Stem Cell 1(1):113-26 PubMed: 18371340
Author